Liver Class‐I Alcohol Dehydrogenase Isozyme Relationships and Constant Patterns in a Variable Basic Structure

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Abstract
The major ethanol dehydrogenase of cobra liver was characterized in order to clarify isozyme relationships and functional motifs of the vertebrate enzyme. The cobra protein is a class-I form, most related to one of the isozyme subunits (the a form) in Uromastix (lizard) liver. This positions the isozyme duplication and defines the main-line alternative. The new structure also allows extensive correlations with structure/function relationships for alcohol dehydrogenases in general, of which 38 animal variants (still disregarding strain and allelic differences) now have been characterized. Architectural features are discerned, distinguishing the enzyme at large, the classes, and the functional interactions at the sites of substrate binding and coenzyme binding. Variability is greater at the substrate-binding site, with only one of 13 residues strictly conserved (His67, one of the active-site zinc ligands) but all other residues differing among and frequently within classes. However, many substrate-interacting residues are class preferential and may be used in predictive assignments. Class-I/III differences concern position 48 (typically Ser in class I, Thr in class III), position 93 (Phe versus Tyr), position 141 (branch-chained aliphatic residue versus methionine), position 57 (hydrophobic residue versus Asp), position 115 (Asp versus Arg), position 116 (Leu or Ile versus Val), position 306 (Met or Leu/Ile versus Phe), position 309 (Phe or Leu/Ile versus Val) and position 318 (Val or Ile versus Ala). In contrast, coenzyme binding is more conserved. A characteristic coenzyme-binging motif, covering only a 50-residue stretch, is defined as tVDiK (residues 178, 203, 223, 224, 228; capital letters for residues strictly conserved and small-cases letters for residues nearly so). This motif is class independent and unique to animal alcohol dehydrogenases. Therefore, the novel enzyme structure establishes class-I isozyme relationships, shows characteristic 'constant' residues also in the 'variable' class-I line, and defines residue-specific patterns which may have a predictive value in functional assignments of an increasing number of undefined further forms expected to result from gene projects.